1. Technical Field of the Invention
The present invention relates to certain isolated polypeptides, to mixtures of polypeptides derived from proteolysis of said isolated polypeptides, to compositions comprised thereof and to cosmetic treatments for reducing intercorneocyte cohesion, and, therefore, for promoting desquamation.
2. Description of the Prior Art
The skin constitutes a physical barrier between the body and its surroundings. It consists of two tissues: the epidermis and the dermis.
The dermis provides the epidermis with a solid support. It is also its feeder component. It consists principally of fibroblasts and an extracellular matrix itself principally composed of collagen, elastin and a substance deemed "ground substance," these components being synthesized by the fibroblast. Leucocytes, mastocytes or tissue macrophages are also found therein. It also comprises blood vessels and nerve fibers.
The epidermis is a desquamative pluristratified epithelium 100 .mu.m thick on average and is conventionally divided into a basal layer of keratinocytes which constitutes the germinative layer of the epidermis, a so-called prickle cell layer consisting of several layers of polyhedral cells arranged on the germinative cells, a so-called granular layer including flattened cells containing distinct cytoplasmic inclusions, the keratohyalin granules, and, finally, a top layer designated the horny layer (or stratum corneum), including keratinocytes at the final stage of their differentiation, designated corneocytes. These are anucleated, mummified cells which are derived from the keratinocytes.
The corneocytes are principally composed of a fibrous matrix containing cytokeratins, surrounded by a very resistant structure 15 nm thick, designated horny or hornified envelope. The stacking of these corneocytes constitutes the horny layer which is responsible for the barrier function of the epidermis.
The stratum corneum possesses a selective permeability which, by controlling the loss of water, ensures a physiological hydration of the skin. It further constitutes a barrier against challenge from or attacks by the surroundings, whether chemical or physical. The stratum corneum is composed of two parts:
(1) the stratum corneum compactum whose cellular organization corresponds to a columnar stacking of the corneocytes above the granular cells from which they are derived. Each corneocyte exhibits maximum covering with the super- and subjacent corneocytes; PA1 (2) the stratum disjunctum including the final strata of the horny layer which are less cohesive than the above and the site of desquamation of the corneocytes. PA1 (i) aspartic acid proteases comprising an aspartic acid residue at the active site thereof, PA1 (ii) serine proteases comprising a serine moiety at the active site thereof, PA1 (iii) cysteine proteases comprising a cysteine moiety at the active site thereof, and PA1 (iv) metalloproteases which most typically comprise a zinc atom at an active site thereof and sometimes a calcium atom. PA1 (a) xerosis (or dryness of the skin), PA1 (b) ichthyoses, PA1 (c) psoriasis, PA1 (d) certain benign or malignant tumor lesions, PA1 (e) reactive hyperkeratoses. PA1 (a) leukokeratosis of the uterine neck during prolapsus, PA1 (b) buccal leukokeratoses, PA1 (c) keratotic benign tumor lesions of the Malpighian mucosae.
In the stratum corneum, the intercorneocyte space is filled with lipid sheets derived from lamellar bodies. Epidermal differentiation represents a continual and oriented maturation process which, from basal keratinocytes results in the formation of corneocytes which are completely keratinized dead cells. This differentiation is the result of perfectly coordinated phenomena which will lead to a constant thickness being maintained and which will thus ensure homeostasis of the epidermis. This passes via a regulation of the number of cells which enter the differentiation process and the number of cells which desquamate.
During the normal desquamation process, only the most superficial corneocytes become detached from the surface of the epidermis.
From the basal layer to the granular layer, cohesion is provided by the transcellular network defined by the desmosomes and the intermediate filaments of cytokeratins. This network is anchored onto the basal membrane by the hemidesmosomes.
In the horny layer, cohesion is provided by the intercellular structures derived from the desmosomes, designated corneosomes or corneodesmosomes, which firmly connect together the horny envelopes of the corneocytes. In the epidermis (non-palmoplantar), the corneodesmosomes are present over the entire corneocyte surface in the lower part of the horny layer, but just the peripheral corneodesmosomes persist in the upper part.
Recent studies have shown the key importance of the corneodesmosomes in intercorneocyte cohesion, as well as in the desquamation process. In particular, a close correlation exists between cell dissociation and proteolysis of certain corneodesmosomal components such as desmoglein I.
The study of desquamation makes it possible to demonstrate the existence of a fine biochemical regulation as far as the so-called "dead" layers of the epidermis. It is the enzymes produced in the deeper living layers which will act sequentially and in a complementary manner, resulting in the final release of the corneocytes at the skin surface.
The principal enzymes suspected of taking part in desquamation have been recently described. They belong to two families of enzymes: the glycosidases and the proteases. The proteases cannot act alone and a preliminary action of glycosidases desquamating sites of proteolysis appears to be necessary.
Proteases constitute the type of enzymes which is probably the most involved in desquamation. They are subdivided into four families:
Among these proteases, the cysteine proteases of lysosomal origin (cathepsins B, H and L) are without doubt the most active proteases in the human body. They are the ones which would participate in the daily renewal of an individual's proteins (from 200 to 300 g for an individual of 70 kg).
In this regard, WO-95/07,686 describes two (2) cysteine proteases of apparent molecular weights 34 to 35 kilodaltons.
Numerous pathological conditions of the skin are characterized by the production of a thick horny layer and by an abnormal desquamation, namely, by hyperkeratbsis. The latter may occur on any anatomical skin area and in a wide variety of clinical contexts. Its physiopathalogical substratum and its cause are varied.
By way of example, representative thereof are:
Other pathological conditions are characterized by transdifferentation or metaplasia, at the level of the mucosae, Malpighian or otherwise, but normally nonhornified, which become hornified, i.e., which become covered with an abnormal epithelium, producing a horny layer at its surface. Although the genital mucosae and those of the upper aerodigestive tracts are most often involved, these metaplasias may be seated in various anatomical areas. Exemplary thereof are:
Without wishing to be bound by or to any particular theory of the invention, it is considered that these pathological conditions may be linked to a qualitative or quantitative deficiency in enzymes suspected of participating in desquamation, including, in particular, proteases.
The purification and recognition of novel polypeptides involved in intercorneocyte cohesion, in particular of proteases, is one of the routes which could allow the production of new species for combating the effects of an excess or a deficiency of polypeptides, in particular of proteases, principally at the surface of the skin or of the mucosae.